Publication Date

2012

Document Type

Dissertation

Committee Members

Steven Berberich (Committee Member), Katherine Excoffon (Committee Member), Michael Leffak (Advisor), Robert Putnam (Committee Chair), Yongjie Xu (Committee Member)

Degree Name

Doctor of Philosophy (PhD)

Abstract

Eukaryotic cells have evolved tightly controlled processes that make sure the entire genome gets replicated accurately, once per cell cycle. This tight control is fulfilled by degrading replication proteins after their function is carried out, changing their cellular location or by post-translational modification of replication factors to regulate their function. Phosphorylation events carried out by CDKs (cyclin dependent kinases) and DDKs (Dbf4 dependent kinases) play crucial roles in this regulation.

The DNA unwinding element binding protein (DUE-B) is an essential replication protein that binds to the human c-myc DNA replication origin. In this study, I find that DUE-B is also actively regulated by kinases and phosphatases. DUE-B binds to chromatin after pre-replication complex formation but before Cdc45 chromatin loading. DUE-B chromatin binding requires CDK activity but it is inhibited by the DNA replication checkpoint response. DDK and PP2A are identified in this study as the primary kinase-phosphatase pair that controls DUE-B phosphorylation. Cdk2 and CKII also phosphorylate DUE-B at the C-terminus domain. Here I show that DUE-B is phosphorylated during the G1-S phase of the cell cycle and gets dephosphorylated in G2-M phases. Phosphorylation on DUE-B controls its function in supporting DNA replication. Specifically, dephosphorylated DUE-B can interact with MCM2-7 complex and inhibit DNA replication. The DUE-B C-terminus domain is very rich in serine and threonine, which can be phosphorylated by many kinases. Here I show that phosphorylation at the C-terminus domain can actively control the protein activity in supporting DNA replication and cell survival.

Page Count

171

Department or Program

Biomedical Sciences

Year Degree Awarded

2012


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